Tissue engineering scaffolds provide the three-dimensional (3-D) geometry and mechanical framework required for regulating cell behavior and facilitating tissue maturation. Unfortunately, most synthetic scaffolds lack the biological recognition motifs required for seeded cell interaction. In order to impart this recognition, synthetic scaffolds should possess appropriate biological functionality. Here, for the first time, we present a comprehensive study of fibronectin (FN) conjugation onto highly porous 3-D poly(carbonate) urethane scaffolds through grafted poly(acrylic acid) spacers on the urethane backbone. Scanning electron microscopy was used to ensure that the porous structures of the scaffolds were preserved throughout the multiple conjugation steps, and Fourier transform infrared spectroscopy was used to monitor the reaction progress. Toluidine blue staining revealed that increasing acrylic acid concentration and grafting time increased the number of poly(acrylic acid) groups incorporated. High resolution X-ray photoelectron spectroscopy studies of the scaffolds demonstrated an increase in nitrogen and sulfur due to FN conjugation. Immunofluorescence microscopy studies showed an even distribution of conjugated FN on the 3-D scaffolds. Cell culture studies using human coronary artery smooth muscle cells demonstrated that FN-conjugated scaffolds had improved cell attachment and infiltration depth compared with scaffolds without FN conjugation and with those scaffolds on which FN was merely adsorbed.